The overall objective of the research proposed in this application is to understand the molecular mechanisms that drive immunologic processes. It has become clear that the events that control lymphoid differentiation are a result of changing patterns of gene expression. The studies proposed here are focused on a class of transcription factors, helix-loop-helix proteins, that are in part responsible for the alterations in gene expression during B cell development. One particular class of helix- loop-helix (HLH) proteins are the E-proteins, that include E12, E47, E2-2 and HEB. E12 and E47 are encoded by one gene, E2A, and arise through differential splicing. E2-2 and HEB are encoded by different genes. The applicant showed previously that E2A gene products are required for proper B cell development. Differentiation of B cells in E2A mutant mice is blocked at a stage prior to immunoglobulin (Ig) gene rearrangement. Some B cell specific transcripts including B29 and Ig germline transcript - are expressed. Others, including RAG-1, RAG-2, mb-1, l5, VpreB and CD19 are absent. In addition, two transcription factors, including Pax -5 and EBF, implicated in the regulation of CD19 and mb-1 respectively, are absent in E2A (-/-) mice. These data suggested that E2A is a central regulator in early B cell development. It is proposed to continue these studies. The specific aims of the are focused on a further understanding of the role of E-proteins in B cell development. Specifically, the applicant plans to examine the individual roles of E12 and E47 in B cell differentiation. He will examine whether E12 or E47 regulate PAX-5 and EBF expression. The individual roles of Pax-5 and EBF in generating the E2A deficient phenotype will be studied. The role of E2-2 in lymphoid development will be investigated using gene targeting. It is planned to use transgenic mice to further dissect the contributions and redundancies of E-proteins in B cell development. Finally, the applicant will perform a genetic analysis of transcription factors in B cell development, including E-proteins, Oct-2 and PU.1.